Project Summary Borrelia burgdorferi is an obligate parasite, and is maintained in nature through a complex cycle involving both a tick and mammalian host. The transition between these two very different host types requires the ability to rapidly adapt through changes in gene expression. Previous studies have provided information on a number of genes that may be differentially expressed under conditions intended to mimic that of the vertebrate or arthropod host. Despite these advances, there remains a fundamental gap in our understanding of the role and function of many genes for host adaptation. Our long-term goals are to identify and characterize the proteins necessary for adaptation of B. burgdorferi to both the tick and mammalian host environments. The overall objective of this application is to identify B. burgdorferi candidate genes required for adapting specifically to the arthropod host. Based on cited studies and preliminary data, the central hypothesis of this proposal is that many B. burgdorferi genes of unknown function are adaptively expressed during uptake and colonization of the tick vector. Furthermore, we hypothesize that some genes important for host adaptation are either transiently expressed, or are induced only during late-stage infection of ticks. The rationale for the proposed research is that once the genes involved in adapting to the tick environment have been identified, it will be possible to determine their respective roles in establishing arthropod infection. This has the potential for important breakthroughs in the field of B. burgdorferi adaptive gene expression that will ultimately lead to strategies to disrupt the enzootic life cycle of the Lyme disease pathogen. Thus, the proposed research is relevant to that part of NIH?s mission that pertains to developing fundamental knowledge that will potentially help to reduce the burdens of human illness and disability. Guided by cited work and preliminary data, our hypothesis will be tested by pursuing two specific aims: 1) Identify genes involved in the adaptation of B. burgdorferi to the arthropod vector environment; and 2) Identify transiently expressed host adaptation factors of B. burgdorferi important for tick infection. Under the first aim, we will make use of In Vivo Expression Technology (IVET). This system will utilize a promoter-based genomic DNA library of B. burgdorferi that is designed to allow for the selection of only those promoters that are active exclusively within the tick host. The second aim will utilize a modified version of the IVET system, termed Recombinase-based IVET (RIVET). This system will also make use of a promoter-based genomic DNA library of B. burgdorferi designed to allow identification of genes important for infection at specific stages of the tick life cycle. When applied, the results from the proposed studies are expected to ultimately lead to new control measures to disrupt the pathogen?s enzootic cycle and prevent infection by the Lyme disease spirochete.